Understanding the renin–angiotensin–aldosterone–SARS-CoV axis: acomprehensive review

Nicholas E. Ingraham 1, Abdo G. Barakat2, Ronald Reilkoff1, Tamara Bezdicek 3,Timothy Schacker4, Jeffrey G. Chipman5, Christopher J. Tignanelli5,6 andMichael A. Puskarich7,8

@ERSpublicationsThe interplay of SARS-CoV-2 with the renin–angiotensin–aldosterone system probably accounts formuch of its unique pathology. Appreciating the degree and mechanism of this interaction highlightspotential therapeutic options, including blockade (ARBs). https://bit.ly/3aue4tS

Cite this article as: Ingraham NE, Barakat AG, Reilkoff R, et al. Understanding the renin–angiotensin–aldosterone–SARS-CoV axis: a comprehensive review. Eur Respir J 2020; 56: 2000912 [https://doi.org/10.1183/13993003.00912-2020].

ABSTRACTImportance: Coronavirus disease 2019 (COVID-19), the disease caused by the severe acute respiratorysyndrome coronavirus 2 (SARS-CoV-2), has been declared a global pandemic with significant morbidityand mortality since first appearing in Wuhan, China, in late 2019. As many countries are grappling withthe onset of their epidemics, pharmacotherapeutics remain lacking. The window of opportunity to mitigatedownstream morbidity and mortality is narrow but remains open. The renin–angiotensin–aldosteronesystem (RAAS) is crucial to the homeostasis of both the cardiovascular and respiratory systems.Importantly, SARS-CoV-2 utilises and interrupts this pathway directly, which could be described as therenin–angiotensin–aldosterone–SARS-CoV (RAAS–SCoV) axis. There exists significant controversy andconfusion surrounding how anti-hypertensive agents might function along this pathway. This reviewexplores the current state of knowledge regarding the RAAS–SCoV axis (informed by prior studies ofSARS-CoV), how this relates to our currently evolving pandemic, and how these insights might guide ournext steps in an evidence-based manner.Observations: This review discusses the role of the RAAS–SCoV axis in acute lung injury and the effects,risks and benefits of pharmacological modification of this axis. There may be an opportunity to leveragethe different aspects of RAAS inhibitors to mitigate indirect viral-induced lung injury. Concerns have beenraised that such modulation might exacerbate the disease. While relevant preclinical, experimental modelsto date favour a protective effect of RAAS–SCoV axis inhibition on both lung injury and survival, clinicaldata related to the role of RAAS modulation in the setting of SARS-CoV-2 remain limited.Conclusion: Proposed interventions for SARS-CoV-2 predominantly focus on viral microbiology and aimto inhibit viral cellular injury. While these therapies are promising, immediate use may not be feasible, andthe time window of their efficacy remains a major unanswered question. An alternative approach is themodulation of the specific downstream pathophysiological effects caused by the virus that lead tomorbidity and mortality. We propose a preponderance of evidence that supports clinical equipoiseregarding the efficacy of RAAS-based interventions, and the imminent need for a multisite randomisedcontrolled clinical trial to evaluate the inhibition of the RAAS–SCoV axis on acute lung injury in COVID-19.

Received: 30 March 2020 | Accepted after revision: 18 April 2020

Copyright ©ERS 2020. This version is distributed under the terms of the Creative Commons Attribution Non-Commercial Licence 4.0.

https://doi.org/10.1183/13993003.00912-2020 Eur Respir J 2020; 56: 2000912

| STATE OF THE ARTINFECTIOUS DISEASE

IntroductionCoronavirus disease 2019 (COVID-19), the infectious disease caused by the severe acute respiratorysyndrome coronavirus 2 (SARS-CoV-2), has left over 180 countries and territories grappling with adevastating pandemic. In December 2019, Wuhan, China, was identified as the epicentre of this outbreak.At the time of writing, reported COVID-19 cases exceeded 700000, with more than 30000 deaths [1–3].While early estimates vary and true values remain uncertain, mortality is estimated between 0.4% and3.4% [4, 5] with initial morbidity and mortality disproportionally affecting older patients [6]. Infectivity(R0) is estimated at 2.24 to 3.58 [4, 7]. COVID-19-related hospitalisations are mostly due to the need forrespiratory support and progressively higher levels of care, with respiratory failure being the underlyingaetiology of COVID-19-related deaths [8, 9]. As many countries are struggling with the onset of theirepidemics, pharmacotherapeutics remain lacking [10, 11]. Learning from prior pandemics and relatedviruses can focus our efforts to control spread and treat those infected [12].

This novel coronavirus is closely related to the severe acute respiratory syndrome coronavirus (SARS-CoV),which caused an outbreak of disease (SARS) in 2003. Genetic studies found that SARS-CoV-2 sharesalmost 80% and 50% sequence identity with SARS-CoV and Middle East respiratory syndromecoronavirus (MERS-CoV), respectively [13]. The genetic variations between SARS-CoV and SARS-CoV-2translate into differences in infectivity [14] and immune response [15]. Despite these differences, theshared genome translates into common clinical, microbiological and biochemical phenotypes.SARS-CoV-2 mimics SARS-CoV’s mechanism of infection, which utilises angiotensin-converting enzyme(ACE) 2, part of the renin–angiotensin–aldosterone system (RAAS). Following virus binding, ACE2activity is downregulated through multiple mechanisms, preventing it from performing its usual functionin states of health, as will be further discussed later in this review [13, 16–18]. The association betweenACE2 and COVID-19 is rooted in two concepts: the mechanism of SARS-CoV-2 infection and theregulatory role of ACE2 during RAAS overactivation. Following the SARS outbreak, extensive researchadvanced our knowledge of highly morbid coronavirus infections [16–25]. The similar renin–angiotensin–aldosterone–SARS-CoV (RAAS–SCoV) axis interactions shared by the two coronaviruses provide anopportunity to further our understanding of this unique interplay in our pursuit of treatment options.

Early attempts to develop safe and effective vaccines for SARS or MERS have been unsuccessful. It ishypothesised that current efforts will take time to develop, and may or may not be efficacious for this orfuture coronavirus pandemics [10, 26–28]. Many potential therapies have been proposed, with a subsetcurrently undergoing investigation [29, 30]. Meanwhile, extensive efforts to date have been appropriatelyfocused on screening, identification and containment, through the collaboration and coordination betweenglobal and local health and governmental agencies [31]. Unfortunately, these public health efforts havehad, at best, mixed success in curbing the spread of the pandemic [32]. Given the rapid spread ofSARS-CoV-2 and the significant morbidity and mortality associated with infection, it behoves the medicalcommunity to evaluate and leverage novel treatments for efficacy, especially if they are already availableand have an established safety profile.

RAAS blockade has been proposed as a potential treatment for SARS-CoV-2 [33–36]. This hypothesis wasinitially published by SUN et al. [35] on 4 February 2020 and reinforced in a publication in DrugDevelopment Research on 4 March 2020 [33]. Other reviews have voiced concern regarding the associationbetween COVID-19 and cardiovascular disease [37], going so far as to postulate that continued RAASblockade may cause harm and to recommend considering discontinuation [38]. The latter argument isbased on the observation that pharmacological blockers of the RAAS can upregulate ACE2 expression,which might increase viral entry into the cell [38]. Evidence from human subjects to support such anassertation is scant, and, as we will see in this review, preclinical and current observational COVID-19evidence would support the contrary hypothesis, that discontinuation of RAAS blockade may proveharmful. These contrasting hypotheses underscore the dire need to evaluate potential mechanisms, if any,through which RAAS modulation would have an impact on the pathophysiology of COVID-19 [35, 37, 39].

Affiliations: 1Dept of Medicine, University of Minnesota, Division of Pulmonary and Critical Care, Minneapolis,MN, USA. 2Dept of Anesthesiology, University of Minnesota, Minneapolis, MN, USA. 3Dept of Pharmacy,Fairview Pharmacy Services, Minneapolis, MN, USA. 4Dept of Medicine, University of Minnesota, Division ofMedicine and Infectious Disease, Minneapolis, MN, USA. 5Dept of Surgery, University of Minnesota, Division ofAcute Care Surgery, Minneapolis, MN, USA. 6Institute for Health Informatics, University of Minnesota,Minneapolis, MN, USA. 7Dept of Emergency Medicine, University of Minnesota, Minneapolis, MN, USA. 8Deptof Emergency Medicine, Hennepin County Medical Center, Minneapolis, MN, USA.

Correspondence: Nicholas E. Ingraham, MMC 276, 420 Delaware St SE, Minneapolis, MN 55455, USA.E-mail: ingra107@umn.edu

https://doi.org/10.1183/13993003.00912-2020 2

INFECTIOUS DISEASE | N.E. INGRAHAM ET AL.

In this review, we intend to compile the existing evidence in order to discuss how we might bridgeknowledge gaps regarding the interplay between SARS-CoV-2, ACE2 and the RAAS.

The RAAS in states of healthOverviewRenin, angiotensin and aldosterone represent the core of a complex hormonal axis, referred to as theRAAS, which contributes to blood pressure control, sodium reabsorption, inflammation and fibrosis [40].RAAS imbalance or modification can cause or treat many diseases, including heart failure, hypotension,diabetes and atherosclerosis [41]. This review focuses on several physiological and pathological effects ofangiotensin II (Ang II) cell signalling (figure 1).

The Ang II/AT1 receptor relationshipAng II, the primary physiological product of the RAAS, is a potent vasoconstrictor. As illustrated in figure1, ACE catalyses the transformation of angiotensin I (Ang I) to Ang II. Ang II elicits its effects byactivating two receptors: the type 1 angiotensin II (AT1) receptor and the type 2 angiotensin II (AT2)receptor [42]. Ang II action through the AT1 receptor causes a cascade with resultant inflammation,vasoconstriction and atherogenesis [43]. These effects also promote insulin resistance and thrombosis [44].In contrast, AT2 receptor stimulation causes vasodilation, decreased platelet aggregation and thepromotion of insulin action. However, the expression of AT2 receptor is low in healthy adults [44]. Thus,Ang II’s effects in adults are modulated and balanced indirectly by ACE2, which converts Ang II intolung-protective Angiotensin-(1–7) (Ang-(1–7)), which gives effects similar to those seen from AT2

receptor stimulation [40, 45]. Conceptually grouping pathways into an Ang II/AT1 receptor pathway andits opposing ACE2/Ang-(1–7) pathway helps with understanding the countering forces and the sequelaewhen an imbalance occurs (figure 1).

The consequences of excessive Ang II include pulmonary vasoconstriction, inflammatory andcytokine-induced organ damage [46] secondary to increased membrane permeability [47], and increasedepithelial cell apoptosis [48]. The effects are predominantly mediated through the unopposed AT1 receptoractivation secondary to decreased ACE2 levels [49]. Furthermore, the pro-inflammatory cascade [50] andincreased vascular permeability [47] caused by over-activation of the AT1 receptor in the lungs directlyinduce acute lung injury and acute respiratory distress syndrome (ARDS), and lead to death [10, 33].

Angiotensin-converting enzyme 2ACE2 is expressed in many tissue types, including respiratory epithelial cells [51]. ACE2 is predominantlyexpressed on the apical surface and converts Ang I and Ang II into lung-protective Ang-(1–9)and Ang-(1–7), respectively, by catalysing a pathway that prevents unopposed AT1 receptor activation.Therefore, ACE2 functions as a counterbalance to its structurally similar counterpart ACE [52], throughthe conversion of Ang II into Ang-(1–7), which, in contrast to ACE, promotes vasodilation, reducesproliferation and prevents apoptosis [52]. ACE2 levels are known to be low in healthy individuals [53] and

Angiotensin II

Angiotensin I

Angiotensinogen

Renin

SARS-CoV-2

Anti-inflammatoryAnti-apoptosisAnti-fibrosis

Pro-inflammatoryPro-apoptosisPro-fibrosis

Spike vaccine

TMPRSS2inhibitor

rhACE2

ARB

ACEi

AT2RAT1R

Ang-(1–7)

ACE ACE2

FIGURE 1 The renin–angiotensin–aldosterone system with COVID-19. The thicker arrows show an increase in the degree of pathway activation;dotted arrows show a decrease in pathway activation. ACE: angiotensin-converting enzyme; ACEi: ACE inhibitors; ARB: angiotensin receptorblocker; AT1R: type 1 angiotensin II receptor; AT2R: type 2 angiotensin II receptor; Ang-(1–7): angiotensin-(1–7); rhACE2: recombinant humanACE2; TMPRSS2: transmembrane serine protease 2.

https://doi.org/10.1183/13993003.00912-2020 3

INFECTIOUS DISEASE | N.E. INGRAHAM ET AL.

ACE2 levels decrease with age [54]. Chronically elevated levels of ACE2 are an independent predictor ofdisease progression, but evidence suggests these changes are compensatory rather than causal [53].

ACE inhibitors and angiotensin receptor blockersBoth ACE inhibitors (ACEi) and angiotensin receptor blockers (ARBs) increase tissue and plasma levels ofrenin and Ang I [55]. ACEi inhibit Ang I to Ang II conversion, decreasing serum Ang II levels to baseline,and have little to no affinity for ACE2 [55]. At clinical doses, ACEi only partially affect this conversionbecause 40% of Ang II formation occurs outside the ACE pathway [56]. ACE levels are in turn decreasedthrough a negative feedback in states of high Ang II levels [57]. Ang II levels increase in response to ARBsat all levels, except in the kidney. The consequence of this increase is a redirection of Ang II activitythrough the anti-inflammatory and vasodilatory ACE2 and AT2 receptor pathways [55]. As a result, ACE2,compared to ACE, regulates local levels of both Ang II and Ang-(1–7) to a greater degree [41]. TIKELLIS

and THOMAS [41] suggest that ARBs, therefore, have a higher potential to favourably rebalance the RAASpathway compared to ACEi. Furthermore, the kallikrein–kinin system is regulated by ACE and ACE2.Increased activity of the kallikrein–kinin system occurs under pro-inflammatory conditions [58],specifically bradykinin and des-Arg9-bradykinin, which can lead to vascular leakage when unchecked byACE and ACE2, respectively. This mechanism has been posited to be, at least in part, a cause of cough[59] and pulmonary oedema [60], independent of proposed Ang II-induced hydrostatic pressure. Thedysregulated kallikrein–kinin system’s role remains theoretical in COVID-19; however, if it is involved,AT1 receptor blockade may be favoured (over ACE inhibition) in an attempt to preserve the regulatorymechanisms of this system [60]. While there may still be a key role for ACE inhibition in the context ofthe current disease, as we will elaborate in the following sections, these observations support our decisionto narrow the focus of this review to the downstream RAAS modifiers, ARBs. A summary of key points isgiven in table 1.

The RAAS in cardiovascular diseaseModulation of the RAAS axis is central to the management of cardiovascular disease [61]. Cardiovasculardisease, heart failure, atrial fibrillation and kidney disease are associated with elevated levels of ACE2 [59].It is critical to discern whether these elevations are a culprit or sequelae of disease in order to separateassociation from causation [62]. More recent opinions suggest that ACE2 represents a biomarker, ratherthan a culprit, of cardiovascular disease [63].

RAMCHAND et al. [53] demonstrated elevated ACE2 levels to be an independent risk factor of major adversecardiac events in patients with known coronary artery disease, although, as noted by the authors, the datado not differentiate association versus causation. To begin to dissect this interplay, we turn to preclinicalmodels, specifically ACE2 knockout mice, which are characterised by severe cardiac defects. Critically,these defects are notably absent in double ACE and ACE2 knockout models, which highlights the highlybalanced interplay between ACE/Ang II/AT1 and ACE2/Ang-(1–7) [52, 64]. Similar protective effects toprevent cardiovascular events and strokes have been demonstrated in mouse models with overexpressedACE2 [65]. RAMCHAND et al. [53] concluded that patients with elevated ACE2 might “reflect a persistentalbeit insufficient counter-regulatory process to shift the balance away from the deleterious effects ofsustained Ang II activation.” As such, ACE2 overexpression may be a compensatory mechanism to mitigatethe effect of unopposed Ang II stimulation through increased degradation into protective Ang-(1–7) [66].

It is important to note that preclinical animal model treatment with losartan upregulates cardiac ACE2mRNA expression and increases ACE2 activity [67, 68]. However, human studies found no significantdifference in ACE2 plasma levels in patients treated with ACEi or ARBs and thus have not confirmed suchan effect, although local tissue ACE2 levels were not measured [53]. Through anecdotal reports,COVID-19 may be associated with high rates of myocarditis and heart failure in the critically ill. While

TABLE 1 Summary of the renin–angiotensin–aldosterone system (RAAS) and its relation toCOVID-19

ACE2 levels are high in diseased states, which is likely to be secondary to an insufficient compensatoryresponse to overactive RAAS activity

In COVID-19, high rates of pulmonary oedema and cough may be due, in part, to reduced breakdown ofbradykinin from decreased ACE activity

ARBs may provide crucial regulation to the overactive RAAS–SCoV axis

ACE: angiotensin-converting enzyme; ARBs: angiotensin receptor blockers; SCoV: SARS-CoV.

https://doi.org/10.1183/13993003.00912-2020 4

INFECTIOUS DISEASE | N.E. INGRAHAM ET AL.

beyond the scope of this article, we will note that increased Ang II and decreased ACE2 are also associatedwith impaired cardiac contractility, which can be rescued with losartan [69]. More observational data areneeded to assess whether other signs of RAAS overdrive (e.g. hypokalaemia or hypertension) portend pooroutcomes and may be associated with arrhythmias that have been noted in the diseases [70]. Nevertheless,this could potentially underscore potential benefits of RAAS modulation that extend beyond theattenuation of lung injury. A further summary of key points is given in table 2.

The RAAS in pulmonary diseaseChronic lung diseaseIn COPD, patients treated with ARBs (compared to those on ACEi) had less severe exacerbations, fewerexacerbations overall, lower mortality, lower mechanical ventilation requirements, and fewer hospitalisations[71]. Moreover, patients aged >65 years who were on ARBs before and during their hospitalisation forpneumonia had decreased mortality when compared to patients without such treatment [72].

Acute lung injuryUnopposed RAAS activation via the Ang II/AT1 receptor pathway causes inflammation [50], increasedvascular permeability [47] and severe lung injury [10, 33], while ARBs significantly attenuate thesechanges [47–50]. Importantly, the mere presence of high concentrations of Ang II can further regulate theexpression of ACE2, leading to dysregulated Ang II/AT1 receptor activity [73]. In mice, losartan reducedmortality by blunting Ang II-associated increases in soluble epoxide hydrolase, a promoter of lung injury[74]. Animal models of ventilator-associated lung injury have demonstrated benefit from losartan,mitigating Ang II activity and AT1 receptor expression [75–77]. While most studies include pre-treatedanimal models, rescue models also demonstrate efficacy, with the restoration of ACE2 levels, blunting ofarterial oxygen tension (PaO2

) decline, and attenuation of lung injury [49].

In human patients, genetic cohort studies yield further insights into the relationship between the RAASand acute lung injury. JERNG et al. [78] found that polymorphisms in the ACE gene are associated withoutcomes in ARDS. These findings were corroborated by ADAMZIK et al. [79], who identified patients withthe ACE DD genotype (associated with increased ACE activity) to have the highest risk of ARDS-relateddeath (hazard ratio 5.7). Other human studies evaluating the association between RAAS inhibition andARDS remain observational. KIM et al. [80] found that ARDS patients that were taking ACEi or ARBs hadbetter survival rates when compared to those without RAAS inhibition. A secondary analysis of a 2010randomised control trial in patients with acute respiratory failure suggested that treatment with ACEi/ARBat discharge following an episode of acute respiratory failure was associated with a 44% reduction in 1-yearmortality [81]. More recently, HSIEH et al. [82] observed lower adjusted odds of hospital mortality inpatients with sepsis (with and without shock) who were on ARB or ACEi therapy. MORTENSEN et al. [83]also found a 58% decrease in the odds of hospital mortality in patients taking ARBs before admission.Based on these data, there have been calls to further elucidate the potential benefits of ACEi and ARBs inARDS [84]. However, randomised control trials around this topic were not identified in the peer-reviewedliterature or ClinicalTrials.gov registry, at the time of writing this article.

PneumoniaWhile influenza and other types of pneumonia may interact with the RAAS axis, both animal and humanstudies illustrate clear indirect effects on the RAAS in the setting of certain influenza strains in particular.Previous studies suggest that Ang II levels predict mortality in undifferentiated patients with influenza[85], and continuation of RAAS inhibitor therapy during admission is associated with decreased hospitalmortality and odds of intubation in viral cases of pneumonia [86]. The RAAS may have implications forother viral pneumonias as well, as GU et al. [87] found that children with respiratory syncytial virus tendto have higher Ang II levels compared to healthy children. Based on this observation, they demonstrated

TABLE 2 Summary of renin–angiotensin–aldosterone system (RAAS) blockade incardiovascular disease states

ACEi and ARBs interrupt maladaptive pathophysiological responses to heightened activation of the RAASARBs and ACEi mitigate the deleterious effects of unopposed AT1 receptor pathway activation, which in

turn decreases inflammation, insulin resistance and lung injury

ACEi: angiotensin-converting enzyme inhibitors; ARBs: angiotensin receptor blockers; AT1: type 1angiotensin II.

https://doi.org/10.1183/13993003.00912-2020 5

INFECTIOUS DISEASE | N.E. INGRAHAM ET AL.

the benefit of recombinant ACE2 therapy on respiratory syncytial virus infection in a preclinical mousemodel.

Critically, both H7N9 and H5N1 influenza have been shown to cause lung injury through ACE2downregulation, upregulation of Ang II, and AT1 receptor-induced lung injury [88, 89]. Mouse models ofH5N1 and H7N9 demonstrated decreased interleukin (IL)-6, lung oedema, lung injury and mortality iftreated with losartan [89, 90]. The mechanism by which losartan prevents lung injury may not residesolely within the RAAS pathway, however [50]. LIU et al. [91] suggest that losartan inhibits the activationof pulmonary dendritic cells. In a study of rats with Pseudomonas pneumonia, AT1 receptor blockerssuppressed the activation of neutrophils through a mechanism that does not involve AT1 receptordownregulation [92]. Such studies have raised concern that losartan treatment might decrease microbialclearance. In sharp contrast, study results actually demonstrate reduced viral loads [90] and increasedbacterial clearance [50] in lung injury models treated with losartan. Given the complexity of theseinteractions, future investigations remain necessary to further elucidate these relationships.

ACE2 binding by SARS-CoVIn 2003, ACE2 was identified as the binding protein for SARS-CoV. In the years to follow, studiesdemonstrating the downstream consequences of downregulation of ACE2 with resultant increases in AngII concentration have illustrated the vicious cycle caused by the disruption of the RAAS, which we refer toin this article as the RAAS–SCoV axis [18]. Of relevance to the current pandemic, WRAPP et al. [14] foundSARS-CoV-2 to have a 10- to 20-fold higher affinity for ACE2 than SARS-CoV. The authors postulate thatthis may cause increased infectivity, which may help explain the differences in the evolution of the twoepidemics.

Both SARS coronaviruses contain four structural proteins: the nucleocapsid, membrane, envelope, andspike protein [93]. The nucleocapsid, membrane and envelope proteins have roles in viral replication,structural integrity and host response, among other mechanisms [93]. The spike protein, known as theS-protein, is integral in attaching to and infecting host cells [14, 93]. PHAN [15] identified specific deletionsthat increase the potential of the SARS-CoV-2 spike to evade immune responses over time. These changeshave implications for both initial and subsequent infection severity, immunity, and the likelihood ofdeveloping an efficacious vaccine. More relevant to the current review, however, is the potential effect suchchanges might have on the RAAS–SCoV axis, which could serve to explain not only their predilection forcausing respiratory failure but possibly other commonalities between these infections.

SARS-CoV infection downregulates the surface expression of the binding protein, ACE2, a pivotalcomponent for host cell entry [17]. Notably, low ACE2 expression is associated with increased phenotypeseverity in human airway epithelial cell in vitro studies [24]. IMAI et al. [23] validated these findings in awhole animal model by demonstrating that loss of ACE2 in knockout mice leads to impairments inoxygenation, increased inflammation, and worsening of oedema compared to wildtype mice afteracid-induced lung injury. Pre-treatment with AT1 receptor inhibitors before acid-induced ARDS showedsignificantly lower lung injury, confirming the AT1 receptor’s role in the physiological response [23].Subsequently and most critically, Kuba et al. [17] demonstrated that SARS-CoV spike protein augmentsAng II increase and ACE2 downregulation, with resultant lung injury. In this model, losartan canattenuate the severity of acute lung injury due to SARS-CoV infection in both a pre-treatment model anda more clinically relevant post-infection (rescue) model [17].

Ensuing studies have confirmed and built upon these findings [19]. AT1 receptor blockade upregulatesACE2 through a negative feedback mechanism, and serves as a lung-protective mechanism through theincreased conversion of Ang II to Ang-(1–7), effectively blunting pulmonary injury from the virus [49].These findings have led investigators to posit that the ACE2/Ang-(1–7) pathway may serve as a therapeutictarget to combat the pathological effects of Ang II [73]. To that end, there is already at least one trial ofrecombinant human ACE2 therapy for the treatment of COVID-19 that attempts to leverage thisrelationship and mitigate downstream lung injury [23, 35].

The mechanism by which SARS-CoV downregulates ACE2 may also be relevant to the development ofnovel therapeutics. HAGA et al. [20] demonstrated that SARS-CoV binding to ACE2 induces ACE2shedding in a soluble form into the serum, and further studies validated those findings [19]. In addition,endocytosis of the ACE2 protein occurs after binding to SARS-CoV, further decreasing ACE2 activity [94],with potential implications for the Ang II/Ang-(1–7) balance. A final critical mechanism that cannot beoverlooked includes the role of the AT1 receptor in the downregulation of ACE2. DESHOTELS et al. [73]found that ACE2 downregulation induced persistent elevation of Ang II through local interaction with theAT1 receptor, which triggers a vicious cycle where Ang II downregulates ACE2 leading to further increasesin local tissue levels of Ang II. These mechanisms promote an unopposed Ang II/AT1 receptor axis (figure 1).

https://doi.org/10.1183/13993003.00912-2020 6

INFECTIOUS DISEASE | N.E. INGRAHAM ET AL.

The ACE2–SARS/COVID-19 interplay has led to controversy with clinical implications. The finding thatACE2 knockout mice are protected against SARS-CoV infection [17] raises a concern that an upregulatedACE2 is harmful in the setting of SARS-CoV and COVID-19 infections, as the virus cannot enter the cell.Conversely, ACE2 downregulation following infection is concerning considering the preclinical datasummarised to this point, which instead suggest a putative causative mechanism of Ang II/Ang-(1–7) inthe evolution of SARS-CoV-2-associated acute lung injury. Even if abolishing ACE2 were feasible, HAN

et al. [21] detected alternative receptors, DC/L-SIGN, which mediated SARS-CoV entry independently ofACE2. A further summary of key points is given in table 3.

Controversies regarding the causative role of ACE2 in COVID-19Limited mechanistic investigations regarding COVID-19 currently exist [78]. Early Chinese data illustrate aprogressive increase in Ang II levels among patients hospitalised in the setting of confirmed COVID-19[95], along with concomitant increasing levels of IL-6, with the highest levels in nonsurvivors [96].Similarly, an analysis in 2014 of H7N9 patients showed progressively increasing levels of Ang II, up to4 weeks following infection, a finding associated with a worse prognosis [89]. In a recent COVID-19patient study, quantified levels of Ang II were positively correlated with viral titres and closely associatedwith PaO2

/inspiratory oxygen fraction measures, suggesting a relationship between infection, the RAAS andlung injury. Accordingly, the study investigators postulated that angiotensin receptor blockade mightrepresent a promising therapeutic target [95].

As noted previously, ACE2 levels are known to be low in healthy individuals [53] and to decrease with age[54]. They are also lower in men than in women [97]. Age is an independent risk factor for mortality inpatients admitted with COVID-19 [96]. The data also suggest a disproportionate rate of diseaseoccurrence, greater severity, and worse outcomes, in males compared to females during this pandemic[98]. These associations might be interpreted to indicate that low ACE2 during times of health areappropriate, but the ability to regulate an overactive RAAS or RAAS–SCoV axis during illness is, at least inpart, dependent on ACE2. For instance, one study identified a higher expression of the lung-protectiveAT2 receptor among women, which could explain the disproportionately lower mortality in femaleCOVID-19 subjects [99]. Interestingly, ACE2 is located on the X chromosome, which has beenhypothesised to affect the ability of men to maintain adequate ACE2 production in situations of severeillness (i.e. ARDS or COVID-19) [39].

Perhaps most importantly, the most common comorbidities of severely ill COVID-19 patients haveincluded hypertension, diabetes mellitus and coronary heart diseases [9]. These comorbidities areassociated with either an overactivation of Ang II/AT1 or a deficiency of ACE2 [41, 44], and all haveshown benefit from RAAS blockade in chronic conditions [100], including an attenuation in AT1 receptorexpression with losartan [101]. Furthermore, smoking tends to increase AT1 receptor expression [101]along with ACE2 expression in human and mouse epithelial cells [102, 103]. RAAS activation in states ofstress with high circulating Ang II levels may be particularly harmful for such patients, and could explainat least some of the observed clinical outcomes, e.g. data from China found that the odds of diseaseprogression in those with COVID-19 are 14 times higher in smokers [8]. However, we acknowledge thatmisinterpreting association for causation can be dangerous and that the pathology underlying acute andchronic conditions may not be equivalent [104]. Nevertheless, RAAS manipulation remains an attractivetherapeutic target, not to treat or prevent infection with COVID-19, but rather to mitigate its downstreamconsequences.

Side-effects of RAAS manipulation are understandably a cause for concern, specifically hypotension andacute kidney injury in the setting of acute illness. The most severe risks of ARBs include teratogenicity inthe setting of pregnancy, hypersensitivity including angioedema, symptomatic hypotension, worsening ofrenal function, and electrolyte abnormalities; however, the most common side-effects include fatigue,

TABLE 3 Summary of renin–angiotensin–aldosterone system interplay with lung injury anddisease

SARS-CoV decreases surface expression of ACE2 during infectionDecreased ACE2 activity leads to increased Ang II and further downregulation of ACE2 in a vicious cycle,driving acute lung injury

While the main point of entry involves ACE2, other receptors can independently mediate SARS-CoVinfection

ACE: angiotensin-converting enzyme; Ang II: angiotensin II.

https://doi.org/10.1183/13993003.00912-2020 7

INFECTIOUS DISEASE | N.E. INGRAHAM ET AL.

weakness, diarrhoea, chest pain and anaemia. Clinical trial and post-marketing surveillance data oflosartan demonstrate an excellent safety profile. In over 4000 patients, there was a low incidence of adverseevents (2.1%) comparable to placebo (3.7%) [105]. The effects more common in losartan than placebowere clinically minor, including dizziness (3% versus 2%), upper respiratory infection (8% versus 7%),nasal congestion (2% versus 1%) and back pain (2% versus 1%). While losartan has the potential to causesymptomatic hypotension, four studies have identified that losartan has a negligible effect on resting bloodpressure and heart rate in healthy volunteers [106–109]. These reassuring data cannot be extrapolated toCOVID-19 patients; however, when assessing the risk/benefit balance for RAAS modulation, the baselinerisk appears low in the healthy population whereas repurposing of other medications may not have thesame favourable profile. There are also concerns that the upregulation of ACE2 by ARBs could accelerateSARS-CoV-2 entry in host cells. Such concerns become more relevant when one considers the observationthat patients with hypertension tend to have worse outcomes, and many patients with hypertension are onagents that might upregulate ACE2 expression. Consequently, some clinicians have argued fordiscontinuing ACEi and ARBs and switching their patients to different anti-hypertensive classes. However,to our knowledge, there are no data to suggest such a change is beneficial, nor are there well-controlled,risk-adjusted analyses demonstrating that these medications are harmful.

On the contrary, the data summarised in this review actually suggest that discontinuation of ACEi orARBs could pose a serious risk of harm through the propagation of excessive Ang II-mediated acute lunginjury and downregulation of protective ACE2. In a recent preprint, LIU et al. [110] retrospectively studied511 COVID-19 patients (age >65 years) with hypertension. Patients were categorised based on homeanti-hypertensive regimen. Patients on ARBs had significantly decreased odds of developing severeCOVID-19 disease on univariate analysis (OR 0.34, p=0.025) and multivariable analysis (OR 0.25,p=0.046), while none of the other medications demonstrated improved outcomes. Two subsequentretrospective studies have demonstrated a protective effect of RAAS blockade, including significantreductions in viral load [111, 112]. Finally, in addition to the inhibitory effects on the AT1 receptor, apreprint publication from Iran identified losartan as a potential candidate treatment based on moleculardocking and dynamic simulation [113]. Therefore, in addition to the effects on angiotensin homeostasis,losartan may directly inhibit the viral macrodomain and cell cycle [113]. Even prior to these observationaldata, based on preclinical evidence and non-COVID-19 studies, the International Society of Hypertension,the European Society of Cardiology and other international committees have strongly recommended thatphysicians and patients continue treatment with their usual anti-hypertensive therapy as there is noclinical or scientific evidence to suggest that treatment with ACEi or ARBs should be discontinued in thesetting of the SARS-CoV-2 pandemic at the current time [114–116].

ConclusionCurrent clinical evidence is insufficient to recommend treating with or withholding RAAS blockade duringthis pandemic. Despite elevated ACE2 levels being found in chronic disease states, the upregulation ofACE2 seems more likely to be an adaptive response rather than a culprit [66]. Significant preclinical dataand retrospective human data suggest RAAS inhibition decreases lung injury and improves survival, whilesimultaneously decreasing viral load in animal models with viral infections that utilise the ACE2 receptor.Human clinical data regarding the effects of RAAS inhibitors on outcomes remain modest and an area ofactive investigation. While the association between hypertension and poor outcomes might be mediatedthrough ACE2 upregulation and increasing viral susceptibility, these differential outcomes are at least aslikely to be a result of dysregulated Ang II-mediated lung injury. We believe this represents a state ofclinical equipoise and hypothesise that RAAS inhibition may improve outcomes in patients infected withCOVID-19. Furthermore, the wide spectrum of severity during the pandemic highlights the heterogeneityin the patients with COVID-19. As different phenotypes emerge, certain subgroups (i.e. those withoveractive RAAS) may differ in their response to therapy. Time will also need to be considered. RAASmodulation may have significantly different effects early in the disease process compared to later, given thecomplexity of the RAAS–SCoV axis. This underscores the exigency for high-quality evidence to investigateany significant association between RAAS modulation and pulmonary pathophysiology, with a focus onACE2-mediated viral infections, particularly SARS-CoV-2.

Acknowledgements: We gratefully acknowledge the editorial assistance of Michael Lotti, Senior Editor of the Universityof Minnesota’s Institute for Engineering in Medicine.

Conflict of interest: None declared.

Support statement: This work was funded by the National Heart, Lung, and Blood Institute, grant T32HL07741.Funding information for this article has been deposited with the Crossref Funder Registry.

https://doi.org/10.1183/13993003.00912-2020 8

INFECTIOUS DISEASE | N.E. INGRAHAM ET AL.

References1 Dong E, Du H, Gardner L. An interactive web-based dashboard to track COVID-19 in real time. Lancet Infect

Dis 2020; 20: 533–534.2 Johns Hopkins University Center for Systems Science and Engineering. Coronavirus COVID-19 (2019-nCoV).

www.arcgis.com/apps/opsdashboard/index.html#/bda7594740fd40299423467b48e9ecf63 World Health Organization. Coronavirus disease (COVID-2019) situation reports. www.who.int/emergencies/

diseases/novel-coronavirus-2019/situation-reports4 Wang Y, Wang Y, Chen Y, et al. Unique epidemiological and clinical features of the emerging 2019 novel

coronavirus pneumonia (COVID-19) implicate special control measures. J Med Virol 2020; 92: 568–576.5 Rajgor DD, Lee MH, Archuleta S, et al. The many estimates of the COVID-19 case fatality rate. Lancet Infect Dis

2020; in press [https://doi.org/10.1016/S1473-3099(20)30244-9].6 WHO-China Joint Mission. Report of the WHO-China Joint Mission on Coronavirus Disease 2019 (COVID-19).

www.who.int/docs/default-source/coronaviruse/who-china-joint-mission-on-covid-19-final-report.pdf Date lastupdated: 24 February 2020.

7 Zhao S, Lin Q, Ran J, et al. Preliminary estimation of the basic reproduction number of novel coronavirus(2019-nCoV) in China, from 2019 to 2020: a data-driven analysis in the early phase of the outbreak. Int J InfectDis 2020; 92: 214–217.

8 Liu W, Tao ZW, Wang L, et al. Analysis of factors associated with disease outcomes in hospitalized patients with2019 novel coronavirus disease. Chin Med J 2020; 133: 1032–1038.

9 Guan WJ, Ni ZY, Hu Y, et al. Clinical characteristics of coronavirus disease 2019 in China. N Engl J Med 2020;382: 1708–1720.

10 Zhang H, Penninger JM, Li Y, et al. Angiotensin-converting enzyme 2 (ACE2) as a SARS-CoV-2 receptor:molecular mechanisms and potential therapeutic target. Intensive Care Med 2020; 46: 586–590.

11 Ingraham NE, Tignanelli CJ. Fact versus science fiction: fighting coronavirus disease 2019 requires the wisdom toknow the difference. Crit Care Explor 2020; 2: e0108.

12 Kuba K, Imai Y, Rao S, et al. Lessons from SARS: control of acute lung failure by the SARS receptor ACE2. JMol Med 2006; 84: 814–820.

13 Zhou P, Yang XL, Wang XG, et al. A pneumonia outbreak associated with a new coronavirus of probable batorigin. Nature 2020; 579: 270–273.

14 Wrapp D, Wang N, Corbett KS, et al. Cryo-EM structure of the 2019-nCoV spike in the prefusion conformation.Science 2020; 367: 1260–1263.

15 Phan T. Genetic diversity and evolution of SARS-CoV-2. Infect Genet Evol 2020; 81: 104260.16 Dijkman R, Jebbink MF, Deijs M, et al. Replication-dependent downregulation of cellular angiotensin-converting

enzyme 2 protein expression by human coronavirus NL63. J Gen Virol 2012; 93: 1924–1929.17 Kuba K, Imai Y, Rao S, et al. A crucial role of angiotensin converting enzyme 2 (ACE2) in SARS

coronavirus-induced lung injury. Nat Med 2005; 11: 875–879.18 Li W, Moore MJ, Vasilieva N, et al. Angiotensin-converting enzyme 2 is a functional receptor for the SARS

coronavirus. Nature 2003; 426: 450–454.19 Glowacka I, Bertram S, Herzog P, et al. Differential downregulation of ACE2 by the spike proteins of severe acute

respiratory syndrome coronavirus and human coronavirus NL63. J Virol 2010; 84: 1198–1205.20 Haga S, Yamamoto N, Nakai-Murakami C, et al. Modulation of TNF-α-converting enzyme by the spike protein

of SARS-CoV and ACE2 induces TNF-α production and facilitates viral entry. Proc Natl Acad Sci USA 2008;105: 7809–7814.

21 Han DP, Lohani M, Cho MW. Specific asparagine-linked glycosylation sites are critical for DC-SIGN- andL-SIGN-mediated severe acute respiratory syndrome coronavirus entry. J Virol 2007; 81: 12029–12039.

22 Hoffmann M, Kleine-Weber H, Schroeder S, et al. SARS-CoV-2 cell entry depends on ACE2 and TMPRSS2 andis blocked by a clinically proven protease inhibitor. Cell 2020; 181: 271–280.e8.

23 Imai Y, Kuba K, Rao S, et al. Angiotensin-converting enzyme 2 protects from severe acute lung failure. Nature2005; 436: 112–116.

24 Jia HP, Look DC, Shi L, et al. ACE2 receptor expression and severe acute respiratory syndrome coronavirusinfection depend on differentiation of human airway epithelia. J Virol 2005; 79: 14614–14621.

25 Jia HP, Look DC, Tan P, et al. Ectodomain shedding of angiotensin converting enzyme 2 in human airwayepithelia. Am J Physiol Lung Cell Mol Physiol 2009; 297: L84–L96.

26 Centers for Disease Control and Prevention. Severe acute respiratory syndrome (SARS) and coronavirus testing –United States, 2003. JAMA 2003; 289: 2203–2206.

27 de Wit E, van Doremalen N, Falzarano D, et al. SARS and MERS: recent insights into emerging coronaviruses.Nat Rev Microbiol 2016; 14: 523–534.

28 Song Z, Xu Y, Bao L, et al. From SARS to MERS, thrusting coronaviruses into the spotlight. Viruses 2019; 11: 59.29 Zhang L, Liu Y. Potential interventions for novel coronavirus in China: a systematic review. J Med Virol 2020; 92:

479–490.30 Li H, Zhou Y, Zhang M, et al. Updated approaches against SARS-CoV-2. Antimicrob Agents Chemother 2020; in

press [https://doi.org/10.1128/AAC.00483-20].31 Fang Y, Nie Y, Penny M. Transmission dynamics of the COVID-19 outbreak and effectiveness of government

interventions: a data-driven analysis. J Med Virol 2020; 92: 645–659.32 Wilder-Smith A, Freedman DO. Isolation, quarantine, social distancing and community containment: pivotal

role for old-style public health measures in the novel coronavirus (2019-nCoV) outbreak. J Travel Med 2020; 27:taaa020.

33 Gurwitz D. Angiotensin receptor blockers as tentative SARS-CoV-2 therapeutics. Drug Dev Res 2020; in press[https://doi.org/10.1002/ddr.21656].

34 Henry B. COVID-19 induced renin–angiotensin system (RAS) imbalance may drive acute lung injury: theevidence and therapeutic options. BMJ 2020; rapid response [www.bmj.com/content/368/bmj.m406/rr-19].

35 Sun ML, Yang JM, Sun YP, et al. [Inhibitors of RAS might be a good choice for the therapy of COVID-19pneumonia]. Zhonghua Jie He He Hu Xi Za Zhi 2020; 43: 219–222.

https://doi.org/10.1183/13993003.00912-2020 9

INFECTIOUS DISEASE | N.E. INGRAHAM ET AL.

36 Tignanelli CJ, Ingraham NE, Sparks MA, et al. Antihypertensive drugs and risk of COVID-19? Lancet RespirMed 2020; 8: e30–e31.

37 Zheng YY, Ma YT, Zhang JY, et al. COVID-19 and the cardiovascular system. Nat Rev Cardiol 2020; 17:259–260.

38 Sommerstein R. Preventing a COVID-19 pandemic: ACE inhibitors as a potential risk factor for fatal COVID-19.BMJ 2020; rapid response [www.bmj.com/content/368/bmj.m810/rr-2].

39 Garami AR. Preventing a COVID-19 pandemic: is there a magic bullet to save COVID-19 patients? We can giveit a try! BMJ 2020; rapid response [www.bmj.com/content/368/bmj.m810/rr-24].

40 Ghazi L, Drawz P. Advances in understanding the renin-angiotensin-aldosterone system (RAAS) in bloodpressure control and recent pivotal trials of RAAS blockade in heart failure and diabetic nephropathy. F1000Res2017; 6: 297.

41 Tikellis C, Thomas MC. Angiotensin-converting enzyme 2 (ACE2) is a key modulator of the renin angiotensinsystem in health and disease. Int J Pept 2012; 2012: 256294.

42 Balakumar P, Jagadeesh G. A century old renin–angiotensin system still grows with endless possibilities: AT1receptor signaling cascades in cardiovascular physiopathology. Cell Signal 2014; 26: 2147–2160.

43 Strawn WB, Ferrario CM. Mechanisms linking angiotensin II and atherogenesis. Curr Opin Lipidol 2002; 13:505–512.

44 Dandona P, Dhindsa S, Ghanim H, et al. Angiotensin II and inflammation: the effect of angiotensin-convertingenzyme inhibition and angiotensin II receptor blockade. J Hum Hypertens 2007; 21: 20–27.

45 Warner FJ, Smith AI, Hooper NM, et al. Angiotensin-converting enzyme-2: a molecular and cellular perspective.Cell Mol Life Sci 2004; 61: 2704–2713.

46 Zhang H, Baker A. Recombinant human ACE2: acing out angiotensin II in ARDS therapy. Crit Care 2017; 21: 305.47 Chen M, Chen C, Yuan X, et al. Angiotensin II aggravates lipopolysaccharide induced human pulmonary

microvascular endothelial cells permeability in high glucose status. Endocr J 2018; 65: 717–725.48 Filippatos G, Uhal BD. Blockade of apoptosis by ACE inhibitors and angiotensin receptor antagonists. Curr

Pharm Des 2003; 9: 707–714.49 Wosten-van Asperen RM, Lutter R, Specht PA, et al. Acute respiratory distress syndrome leads to reduced ratio

of ACE/ACE2 activities and is prevented by angiotensin-(1-7) or an angiotensin II receptor antagonist. J Pathol2011; 225: 618–627.

50 Raiden S, Nahmod K, Nahmod V, et al. Nonpeptide antagonists of AT1 receptor for angiotensin II delay theonset of acute respiratory distress syndrome. J Pharmacol Exp Ther 2002; 303: 45–51.

51 Mossel EC, Wang J, Jeffers S, et al. SARS-CoV replicates in primary human alveolar type II cell cultures but notin type I-like cells. Virology 2008; 372: 127–135.

52 Turner AJ, Hiscox JA, Hooper NM. ACE2: from vasopeptidase to SARS virus receptor. Trends Pharmacol Sci2004; 25: 291–294.

53 Ramchand J, Patel SK, Srivastava PM, et al. Elevated plasma angiotensin converting enzyme 2 activity is anindependent predictor of major adverse cardiac events in patients with obstructive coronary artery disease. PLoSOne 2018; 13: e0198144.

54 AlGhatrif M, Cingolani O, Lakatta EG. The dilemma of coronavirus disease 2019, aging, and cardiovasculardisease: insights from cardiovascular aging science. JAMA Cardiol 2020; in press [https://doi.org/10.1001/jamacardio.2020.1329].

55 Campbell DJ. Endogenous angiotensin II levels and the mechanism of action of angiotensin-converting enzymeinhibitors and angiotensin receptor type 1 antagonists. Clin Exp Pharmacol Physiol Suppl 1996; 3: S125–S131.

56 Hollenberg NK, Fisher ND, Price DA. Pathways for angiotensin II generation in intact human tissue: evidencefrom comparative pharmacological interruption of the renin system. Hypertension 1998; 32: 387–392.

57 Schunkert H, Ingelfinger JR, Hirsch AT, et al. Feedback regulation of angiotensin converting enzyme activity andmRNA levels by angiotensin II. Circ Res 1993; 72: 312–318.

58 Jurado-Palomo J, Caballero T. Pathophysiology of bradykinin-mediated angioedema: the role of the complementsystem. In: Kartal SP, Kutlubay Z, eds. A Comprehensive Review of Urticaria and Angioedema. Rijeka,IntechOpen, 2017; pp. 151–176.

59 Bahat G. Angiotensin converting enzyme (ACE) inhibition may have a role in the symptoms and progression ofCOVID-19 infection. BMJ 2020; rapid response [www.bmj.com/content/368/bmj.m406/rr-23].

60 van de Veerdonk F, Netea MG, van Deuren M, et al. Kinins and cytokines in COVID-19: a comprehensivepathophysiological approach. Preprints 2020; preprint [https://doi.org/10.20944/preprints202004.0023.v1].

61 Braun-Menendez E, Fasciolo JC, Leloir LF, et al. The substance causing renal hypertension. J Physiol 1940; 98:283–298.

62 Walters TE, Kalman JM, Patel SK, et al. Angiotensin converting enzyme 2 activity and human atrial fibrillation:increased plasma angiotensin converting enzyme 2 activity is associated with atrial fibrillation and moreadvanced left atrial structural remodelling. Europace 2017; 19: 1280–1287.

63 Turner AJ. ACE2 cell biology, regulation, and physiological functions. In: Unger T, Steckelings UM, dos SantosRAS, eds. The Protective Arm of the Renin Angiotensin System (RAS). Boston, Academic Press, 2015; pp.185–189.

64 Crackower MA, Sarao R, Oudit GY, et al. Angiotensin-converting enzyme 2 is an essential regulator of heartfunction. Nature 2002; 417: 822–828.

65 Rentzsch B, Todiras M, Iliescu R, et al. Transgenic angiotensin-converting enzyme 2 overexpression in vessels ofSHRSP rats reduces blood pressure and improves endothelial function. Hypertension 2008; 52: 967–973.

66 Turner AJ. Angiotensin-converting enzyme 2: cardioprotective player in the renin-angiotensin system?Hypertension 2008; 52: 816–817.

67 Ferrario CM, Jessup J, Chappell MC, et al. Effect of angiotensin-converting enzyme inhibition and angiotensin IIreceptor blockers on cardiac angiotensin-converting enzyme 2. Circulation 2005; 111: 2605–2610.

68 Ocaranza MP, Godoy I, Jalil JE, et al. Enalapril attenuates downregulation of angiotensin-converting enzyme 2 inthe late phase of ventricular dysfunction in myocardial infarcted rat. Hypertension 2006; 48: 572–578.

69 Perlot T, Penninger JM. ACE2 – from the renin-angiotensin system to gut microbiota and malnutrition. MicrobesInfect 2013; 15: 866–873.

https://doi.org/10.1183/13993003.00912-2020 10

INFECTIOUS DISEASE | N.E. INGRAHAM ET AL.

70 Chen D, Li X, Song Q, et al. Hypokalemia and clinical implications in patients with coronavirus disease 2019(COVID-19). medRxiv 2020; preprint [https://doi.org/10.1101/2020.02.27.20028530].

71 Lai CC, Wang YH, Wang CY, et al. Comparative effects of angiotensin-converting enzyme inhibitors andangiotensin II receptor blockers on the risk of pneumonia and severe exacerbations in patients with COPD. Int JChron Obstruct Pulmon Dis 2018; 13: 867–874.

72 Mortensen EM, Nakashima B, Cornell J, et al. Population-based study of statins, angiotensin II receptorblockers, and angiotensin-converting enzyme inhibitors on pneumonia-related outcomes. Clin Infect Dis 2012; 55:1466–1473.

73 Deshotels MR, Xia H, Sriramula S, et al. Angiotensin II mediates angiotensin converting enzyme type 2internalization and degradation through an angiotensin II type I receptor-dependent mechanism. Hypertension2014; 64: 1368–1375.

74 Tao W, Li PS, Xu G, et al. Soluble epoxide hydrolase plays a vital role in angiotensin II-induced lung injury inmice. Shock 2018; 50: 589–594.

75 Chen C, Zhang Z, Li Z, et al. Losartan attenuates microvascular permeability in mechanical ventilator-inducedlung injury in diabetic mice. Mol Biol Rep 2014; 41: 809–814.

76 Yao S, Feng D, Wu Q, et al. Losartan attenuates ventilator-induced lung injury. J Surg Res 2008; 145: 25–32.77 Ling X, Lou A, Li Y, et al. [Losartan regulates oxidative stress via caveolin-1 and NOX4 in mice with

ventilator-induced lung injury]. Nan Fang Yi Ke Da Xue Xue Bao 2015; 35: 1739–1744.78 Jerng JS, Yu CJ, Wang HC, et al. Polymorphism of the angiotensin-converting enzyme gene affects the outcome

of acute respiratory distress syndrome. Crit Care Med 2006; 34: 1001–1006.79 Adamzik M, Frey U, Sixt S, et al. ACE I/D but not AGT (−6) A/G polymorphism is a risk factor for mortality in

ARDS. Eur Respir J 2007; 29: 482–488.80 Kim J, Choi SM, Lee J, et al. Effect of renin-angiotensin system blockage in patients with acute respiratory

distress syndrome: a retrospective case control study. Korean J Crit Care Med 2017; 32: 154–163.81 Noveanu M, Breidthardt T, Reichlin T, et al. Effect of oral beta-blocker on short and long-term mortality in

patients with acute respiratory failure: results from the BASEL-II-ICU study. Crit Care 2010; 14: R198.82 Hsieh MS, How CK, Hsieh VCR, et al. Preadmission antihypertensive drug use and sepsis outcome: impact of

angiotensin-converting enzyme inhibitors (ACEIs) and angiotensin receptor blockers (ARBs). Shock 2020; 53:407–415.

83 Mortensen EM, Restrepo MI, Copeland LA, et al. Impact of previous statin and angiotensin II receptor blockeruse on mortality in patients hospitalized with sepsis. Pharmacotherapy 2007; 27: 1619–1626.

84 Boyle AJ, Mac Sweeney R, McAuley DF. Pharmacological treatments in ARDS; a state-of-the-art update. BMCMed 2013; 11: 166.

85 Huang F, Guo J, Zou Z, et al. Angiotensin II plasma levels are linked to disease severity and predict fataloutcomes in H7N9-infected patients. Nat Commun 2014; 5: 3595.

86 Henry C, Zaizafoun M, Stock E, et al. Impact of angiotensin-converting enzyme inhibitors and statins on viralpneumonia. Proc (Bayl Univ Med Cent) 2018; 31: 419–423.

87 Gu H, Xie Z, Li T, et al. Angiotensin-converting enzyme 2 inhibits lung injury induced by respiratory syncytialvirus. Sci Rep 2016; 6: 19840.

88 Zou Z, Yan Y, Shu Y, et al. Angiotensin-converting enzyme 2 protects from lethal avian influenza A H5N1infections. Nat Commun 2014; 5: 3594.

89 Yang P, Gu H, Zhao Z, et al. Angiotensin-converting enzyme 2 (ACE2) mediates influenza H7N9 virus-inducedacute lung injury. Sci Rep 2014; 4: 7027.

90 Yan Y, Liu Q, Li N, et al. Angiotensin II receptor blocker as a novel therapy in acute lung injury induced byavian influenza A H5N1 virus infection in mouse. Sci China Life Sci 2015; 58: 208–211.

91 Liu J, Zhang P, Yu T, et al. [Effects of losartan on pulmonary dendritic cells in lipopolysaccharide-induced acutelung injury mice]. Zhonghua Yi Xue Za Zhi 2014; 94: 3216–3219.

92 Raiden S, Pereyra Y, Nahmod V, et al. Losartan, a selective inhibitor of subtype AT1 receptors for angiotensin II,inhibits neutrophil recruitment in the lung triggered by fMLP. J Leukoc Biol 2000; 68: 700–706.

93 Schoeman D, Fielding BC. Coronavirus envelope protein: current knowledge. Virol J 2019; 16: 69.94 Wang S, Guo F, Liu K, et al. Endocytosis of the receptor-binding domain of SARS-CoV spike protein together

with virus receptor ACE2. Virus Res 2008; 136: 8–15.95 Liu Y, Yang Y, Zhang C, et al. Clinical and biochemical indexes from 2019-nCoV infected patients linked to viral

loads and lung injury. Sci China Life Sci 2020; 63: 364–374.96 Zhou F, Yu T, Du R, et al. Clinical course and risk factors for mortality of adult inpatients with COVID-19 in

Wuhan, China: a retrospective cohort study. Lancet 2020; 395: 1054–1062.97 Xudong X, Junzhu C, Xingxiang W, et al. Age- and gender-related difference of ACE2 expression in rat lung. Life

Sci 2006; 78: 2166–2171.98 Cai H. Sex difference and smoking predisposition in patients with COVID-19. Lancet Respir Med 2020; 8: e20.99 Hilliard LM, Jones ES, Steckelings UM, et al. Sex-specific influence of angiotensin type 2 receptor stimulation on

renal function: a novel therapeutic target for hypertension. Hypertension 2012; 59: 409–414.100 Packer M, McMurray JJV. Importance of endogenous compensatory vasoactive peptides in broadening the

effects of inhibitors of the renin-angiotensin system for the treatment of heart failure. Lancet 2017; 389:1831–1840.

101 Podowski M, Calvi C, Metzger S, et al. Angiotensin receptor blockade attenuates cigarette smoke-induced lunginjury and rescues lung architecture in mice. J Clin Invest 2012; 122: 229–240.

102 Leung JM, Yang CX, Tam A, et al. ACE-2 expression in the small airway epithelia of smokers and COPDpatients: implications for COVID-19. Eur Respir J 2020; 55: 2000688.

103 Yilin Z, Yandong N, Faguang J. Role of angiotensin-converting enzyme (ACE) and ACE2 in a rat model ofsmoke inhalation induced acute respiratory distress syndrome. Burns 2015; 41: 1468–1477.

104 Fang L, Karakiulakis G, Roth M. Are patients with hypertension and diabetes mellitus at increased risk forCOVID-19 infection? Lancet Respir Med 2020; 8: e21.

105 Burrell LM. A risk-benefit assessment of losartan potassium in the treatment of hypertension. Drug Saf 1997; 16:56–65.

https://doi.org/10.1183/13993003.00912-2020 11

INFECTIOUS DISEASE | N.E. INGRAHAM ET AL.

106 Christen Y, Waeber B, Nussberger J, et al. Oral administration of DuP 753, a specific angiotensin II receptorantagonist, to normal male volunteers. Inhibition of pressor response to exogenous angiotensin I and II.Circulation 1991; 83: 1333–1342.

107 Doig JK, MacFadyen RJ, Sweet CS, et al. Haemodynamic and renal responses to oral losartan potassium duringsalt depletion or salt repletion in normal human volunteers. J Cardiovasc Pharmacol 1995; 25: 511–517.

108 Ohtawa M, Takayama F, Saitoh K, et al. Pharmacokinetics and biochemical efficacy after single and multiple oraladministration of losartan, an orally active nonpeptide angiotensin II receptor antagonist, in humans. Br J ClinPharmacol 1993; 35: 290–297.

109 Reid JL. Inhibitors of the renin-angiotensin system. Clinical pharmacology studies on kinetics, dynamics andconcentration-effect relationships. Arzneimittelforschung 1993; 43: 263–264.

110 Liu Y, Huang F, Xu J, et al. Anti-hypertensive angiotensin II receptor blockers associated to mitigation of diseaseseverity in elderly COVID-19 patients. medRxiv 2020; preprint [https://doi.org/10.1101/2020.03.20.20039586].

111 Meng J, Xiao G, Zhang J, et al. Renin-angiotensin system inhibitors improve the clinical outcomes of COVID-19patients with hypertension. Emerg Microbes Infect 2020; 9: 757–760.

112 Bean DM, Kraljevic Z, Searle T, et al. Treatment with ACE-inhibitors is associated with less severe disease withSARS-COVID-19 infection in a multi-site UK acute Hospital Trust. medRxiv 2020; preprint [https://doi.org/10.1101/2020.04.07.20056788].

113 Sallari Jazzi A, Mahnam K, Hossein Hejazi S, et al. Inhibition of viral macrodomain of COVID-19 and humanTRPM2 by losartan. Preprints 2020; preprint [https://doi.org/10.20944/preprints202003.0457.v1].

114 International Society of Hypertension. A statement from the International Society of Hypertension onCOVID-19. https://ish-world.com/news/a/A-statement-from-the-International-Society-of-Hypertension-on-COVID-19/ Date last accessed: 30 March 2020.

115 Hypertension Canada. Hypertension, ACE-inhibitors and angiotensin receptor blockers and COVID-19. Markham,Hypertension Canada, 2020. https://hypertension.ca/wp-content/uploads/2020/03/2020-30-15-Hypertension-Canada-Statement-on-COVID-19-ACEi-ARB.pdf

116 Canadian Cardiovascular Society. COVID-19 and concerns regarding use of ACEi/ARB/ARNi medications forheart failure or hypertension. Ottawa, Canadian Cardiovascular Society, 2020. www.conferenceharvester.com/Uploads/Documents/9006/CCSCHFSstatementregardingCOVID.pdf

https://doi.org/10.1183/13993003.00912-2020 12

INFECTIOUS DISEASE | N.E. INGRAHAM ET AL.